MicroRNAs (miRNAs) represent a relatively new family of regulatory RNAs with broad potential revelevance in post-transcritional regulation of developmental and disease pathways. To provide insight into the broader potential of miRNA activities, we propose a series of experiments to investigate the neuralspecific Drosophila miRNA-315. Using co-expression analysis with established neuronal and glial cell markers, we will further characterize the embryonic neural cell subtype(s) which expresses miR-315, focusing specifically on potential roles in regulation of neural stem cell development. We will complement this with in vivo promoter/enhancer analysis to better understand the cis-regulatory control that directs the neural-restricted expression patterns that have been observed, allowing us to better understand the regulatory networks that impinge on miR-315 activity. To increase our knowledge of the role that miRNAs play in target regulation, we will also assess the degree to which miR-315 is co-expressed with computationally-predicted targets. These data will contribute to a more refined model of the generalizable role of miRNAs in post-transcriptional gene regulation. Using these data as a guide for phenotypic characterization, we will approach miR-315 gain and loss of function studies to determine its endogenous function in Drosophila embryonic development. We will examine phenotypes for evidence of neural patterning and morphology defects in addition to testing for rescue by modifying dosages of relevant miR- 315 targets as well as re-introduction of miR-315 expression in mutant embryos. To guide our efforts, we will use in vitro sensor assays to further refine our understanding of the transcripts representing true miR- 315 targets, thereby providing a clearer picture of the relevant regulatory capacity of this miRNA. Collectively, these data will provide one of the first examples of endogenous miRNA function in the developing nervous system. There is already strong emerging evidence for miRNAs playing important roles as potential oncogenes in cancer, as well as demonstrating activity in immmunopathologies, neurodegenerative disease and stem cell maintenance. The majority of these studies rely solely on the impact of miRNA misexpression or loss of global miRNA processing. In addition to extensive misexpression analysis, our study will address the developmental consequences of specific miRNA loss of function, information that we expect will contribute to our general understanding of how miRNAs integrate into regulatory networks controlling neurogenesis and how perturbation of these activites through aberrant expression might contribute to neurodegenerative diseases and other relevant human pathologies.